Abstract Poly-ADP-ribosylation (PARylation) is a protein posttranslational modification (PTM) that was first documented in 1963. It is catalyzed by a family of enzymes called Poly-ADP-ribose polymerases (PARPs). PARP1 is a nuclear enzyme that is involved in cell stress responses. In particular, the critical roles of PARP1 in mediating DNA damage response (DDR) provide the rationale for developing PARP1 inhibitors for the treatment of cancer. Besides regulating DDR in the context of human malignancies, poly-ADP-ribose is known to be a death signal, and PARP1 has been recently proposed as a promising therapeutic target for neurodegenerative diseases. The mechanism by which aberrant PARP1 activation causes neuronal cell death, however, is incompletely understood. Very few genuine PARP1 substrates have been identified, and in most of the cases, how PARylation regulates the function of these potential cell death effectors is poorly defined. Protein PARylation represents a daunting challenge for mass spectrometrists, due to its low-abundance, labile and heterogeneous nature. Recently, we developed the first large-scale mass spectrometric (MS) approach towards site-specific characterization of the Asp- and Glu-PARylated proteome. For the current proposal (R01 supplements), we will leverage these preliminary data, to profile the PARylated proteome that is relevant to neurodegenerative diseases. We will perform quantitative MS characterization, in neurons, of the pathologic PARylated proteome induced by various neurotoxins, and generate a comprehensive map for protein PARylation for neurodegenerative diseases. We expect that these results will serve as an invaluable resource, providing the foundation for future hypothesis-driven research that helps delineate the molecular mechanisms by which PARP1 regulates the pathogenesis of these devastating diseases.